Objective To find a feasible method that can reconstruct the composite tissue-engineered skin fast in vitro and can provide enoughskin as soon as possible for covering the surface of the large-area burn. Methods The foreskin was taken during the posthectomy. The epidermal cells and fibroblasts were isolated, identified and cultured. The cytokeratin 19 (K19) flow cytometry and the fluorescein isothiocyanate (FITC)immunofluorescence for K19 and the FITCimmunofluorescence for PAN-cytokeratin of the epidermal cells and the FITCimmunofluorescence for vimentin of the fibroblasts were performed to identify the epidermal cells and the fibroblasts. Then, the epidermal cells were seeded onto the papillary surface of an acellular dermal matrix (ADM) and were submerged into the condition culture medium added with 25 ng/ml of the keratinocyte growth factor (KGF). However, in the control group, no KGFwas added. After 24 hours, the ADM was moved up to the airfluid surface, and the culture was continued. After 6 days, the fibroblasts were seeded onto the other surface of the ADM. After a 24 hour culture, the ADM was harvested and fixed in formalin, and the hematoxylin-eosin staining was conducted. Then, the structure of the reconstructed skin was observed under the microscope and the cell count in the epidermal layer was also conducted. Results All the cultured and expanded epidermal cells stained by the immunofluorescence demonstrated a positive reaction for PANFITC, and a partially positive for K19-FITC, and 17% of the cells demonstrated a positive reaction for K19 identified by the flow cytometry. The fibroblasts could be expanded by more than 100 times after a 7day culture in vitro, and they could demonstrate a positive reaction for vimentin-FITC. After a 7day culture, a composite tissue-engineered skin could be attained. The hematoxylineosin staining of the reconstructed skin showed that there was one continuous layer of the epidermis on the papillary surface of the ADM and there were fibroblasts in the superficial layer of the other one, but the epidermal layer did not stick tightly to the ADM. The cell count demonstrated that KGF promoted the epidermal cells to proliferate better（Plt;0.01）and to form more significantly continuous layers of the epidermis in the experimental group than in the control group（Plt;0.01）. Conclusion Through the seed-cell separation by the digestion of collagenase and trypsin combined with the use of the KGF-added condition ulture medium, a composite tissueengineered skin can be reconstructed within 7 days.
Objective To review the research progress in the construction strategy and application of bone/cartilage immunomodulating hydrogels. Methods The literature related to bone/cartilage immunomodulating hydrogels at home and abroad in recent years was reviewed and summarized from the immune response mechanism of different immune cells, the construction strategy of immunomodulating hydrogels, and their practical applications. Results According to the immune response mechanism of different immune cells, the biological materials with immunoregulatory effect is designed, which can regulate the immune response of the body and thus promote the regeneration of bone/cartilage tissue. Immunomodulating hydrogels have good biocompatibility, adjustability, and multifunctionality. By regulating the physical and chemical properties of hydrogel and loading factors or cells, the immune system of the body can be purposively regulated, thus forming an immune microenvironment conducive to osteochondral regeneration. ConclusionImmunomodulating hydrogels can promote osteochondral repair by affecting the immunomodulation process of host organs or cells. It has shown a wide application prospect in the repair of osteochondral defects. However, more data support from basic and clinical experiments is needed for this material to further advance its clinical translation process.
目的 探讨成都市传染病医院护理应急体系的构建方法、效果。 方法 成立护理应急管理小组；组建护理应急梯队；储备应急物资和设备；加强护理应急人员知识技能培训和实战演练；严格防护措施与消毒隔离流程。 结果 出色地完成了多次突发传染病的救治工作，培养了一支具有丰富应急救治经验的专业护理人员队伍。 结论 建立完善的护理应急体系可有效提高突发事件的应急保障能力。
This study introduced the construction of individualized risk assessment model based on Bayesian networks, comparing with traditional regression-based logistic models using practical examples. It evaluates the model's performance and demonstrates its implementation in the R software, serving as a valuable reference for researchers seeking to understand and utilize Bayesian network models.
With the development of tissue engineering, a variety of forms of silk fibroin (SF) scaffolds has been applied to research of constructing variety of organization based on cells, which has become scientific focus in recent years. In this paper we introduced the source and structure of SF and the fabrication method of the scaffold, and also address the SF application progress in several relevant fields of tissue engineering, such as bone, cartilage, skin, blood vessel and nerves. Finally, we discuss the future leading prospect of the SF in order to provide reference for subsequent research.
ObjectiveTo evaluate the developing methodologies of Essential Medicines Lists for Children (EMLcs) in global, in order to provide reference in developing EMLc of China. MethodsWe searched ProQuest, ScienceDirect, SpringerLink and MEDLINE databases, World Health Organization (WHO) official website, and 67 websites of National Ministry of Health and Drug Administration Section, to collect literature about selection methodology of children and/or adult essential medicines list (EML). A descriptive analysis was conducted. ResultsA total of fourteen literatures were included. Of which, 6 were about the essential medicines selection methodology in children, and the other 8 were about the essential medicines selection methodology in adult. The WHO had established independent EMLc selection committee. Paediatricians were involved in the selection of EMLc in the WHO and India. There was no selection criteria and process for EMLc globally. The WHO, India, and South Africa selected their EMLcs referring to the WHO EML selection criteria. The WHO and South Africa had their own updating time, period and process for EMLc. The WHO EMLc was updated per 2 years, which in high frequency and conducts in rigorous process. However, the EMLc of India had not been updated yet. ConclusionIt is suggested that China could build a national EMLc selection committee involving paediatricians and evidence-based medicine experts etc. in referring to the framework of the WHO Child Health Working Group. The EMLc selection criteria and process of China could be established referring to the one of the WHO, based on the disease burden, drug accessibility and medical insurance of children of China. The EMLc of China should be simultaneously updated with the adult EML of China.
Objective To search the most suitable concentration of calcium in the medium for the basement membrane reconstruction in tissue engineering skin in vitro. Methods Composite chitosan tissue engineering skin was prepared according to previous studies. Four groups were included according to the concentrationof calcium (1.00, 1.45, 1.65 and 1.95 mmol/L respectively). After 7 days and 15 days of culture, the histological manifestation of basement membrane in tissue engineering skin was observed by hematoxylin amp; eosin staining and PAS staining, and collagen Ⅳ of basement membrane was detected immunohistochemicallyatthe dermalepidermal junction. Results This tissue engineering skin shared some histological features of normal skin, including a welldifferentiated stratifiedepidermis and a dense dermis. The epithelium in the group of 1.95 mmol/L calcium differentiated better than those in other groups. PAS staining showing a regularly red dying strap domain at the dermal-epidermal junction. Collagen Ⅳ was positively stained immunohistochemically at the dermalepidermal junction inthe tissue engineering skin. Conclusion The above results suggest that the medium with 1.95 mmol/L calcium should be suitable for the growth of composite chitosan tissue engineering skin and the reconstruction of basement membrane.
Objective To construct yeast eukaryotic expression vector carrying human endostatin (ES) cDNA. Methods The functional fragment of endostatin gene in human hepatic tissue was amplified by using RT-PCR technology, and cloned into yeast pPIC9 expression vector. The positive clone was sequenced by using automatized sequencer. Results The endostatin cDNA was successfully cloned. The positive ES clone gene in pPIC9 expression vector was sieved, and its coding sequence was identified to be as same as the previously reported sequence. Conclusion The successful construction of ES gene in pPIC9 expression vector using molecular biological method maybe helpful for the high expression of ES protein, which may lay the foundation for the treatment of malignant tumor through anti-angiogenesis appoach.
Objective To construct gene-modified hepatic stem cells (WB-F344 cells), which have rat IL-13 gene and can secrete the recombinant rat IL-13 cytokine in the cells. Methods Firstly, the rat IL-13 sequences were synthesized. Then the sequences were amplificated in bacterium coli after recombinated with pWPXL-MOD plasmid. After PCR and sequence identification, the positive clones were packaged into lentivirus. After detecting the virus titer, the WB-F344 cells with constructed lentivirus vector with rat IL-13 gene were cultured, then the valid targets (expression level of the IL-13) were detected by real time-PCR and Western blot in cultured WB-F344 cells on 5 days. Results The valid DNA of rat IL-13 was recombinated and packaged in lentivirus vector. The recombinant gene sequence was correct by checking with gene sequence test. Then the recombinant was introducted into the WB-F344 cells cultures. The best multiplicity of infection (MOI) value for effective transfection was 5. IL-13 had been detected on day 5 after transfection by checking with real-time PCR and Western blot. Conclusion The recombinant rat IL-13 gene with lentivirus vector is constructed and gene-modified WB-F344 cells are cultured successfully, which can be used in next animal experiment.
Objective To construct and screen neurite outgrowth inhibitory 66-samll interfering RNA (nogo66-siRNA) eukaryotic expression vectors of effective interference, so as to lay a foundation for further reconstruction of related viral vector. Methods The nogo66-siRNA fragments were designed and cloned into pGenesil-1.1, 4 plasmids of pGenesil-nogo66-siRNA-1, pGenesil-nogo66-siRNA-2, pGenesil-nogo66-siRNA-hk, and pGenesil-nogo66-siRNA-kb were obtained, sequenced and identified, then were transfected into C6 cell l ine. The transfection efficiency was measured by fluorescence microscope. RT-PCR and Western blot were used to detect the expression of nogo gene and select the plasmid of effective interference. Results DNA sequencing results showed interference sequences were correct. The bands of 800 bp and 4.3 kb were detected when pGenesil-nogo66-siRNAs were digested by Kpn I /Xho I. The expression of green fluorescent protein could be detected under fluorescence microscope, and the transfection efficiency was about 73%. RT-PCR and Western blot results showed that compared to non-transfected cells, the transfection of pGenesil-nogo66-siRNA-1 made the expression of nogo gene decl ine 22% and the expression of nogo protein decl ine 73%; the transfection of pGenesil-nogo66-siRNA-2 made the expression of nogo gene decl ine 28% and the expression of nogo protein decl ine 78%; the differences were significant (P lt; 0.05); and the transfection of pGenesil-nogo66-siRNA-hk and pGenesil-nogo66- siRNA-kb did not make the expressions of nogo gene and nogo protein decrease significantly (P gt; 0.05). Conclusion Nogo66-siRNA eukaryotic expression vector is successfully constructed, it lays an experimental foundation for repair of spinal cord injury.